Atg21 Is Required for Effective Recruitment of Atg8 to the Preautophagosomal Structure during the Cvt Pathway

Abstract

Atg21 and Atg18 are homologue yeast proteins. Whereas Atg18 is essential for the Cvt pathway and autophagy, a lack of Atg21 only blocks the Cvt pathway. Our proteinase protection experiments now demonstrate that growing atg21Δ cells fail to form proaminopeptidase I-containing Cvt vesicles. Quantitative measurement of autophagy atg21Δ cells only of the that Atg21 a the of of Atg21 is the proteins. cells to the of the a of the is to the and growing Atg21 is for of to the the of to the is is to the of the and the of of atg21Δ cells the of of Atg21 and Atg18 are homologue yeast proteins. Whereas Atg18 is essential for the Cvt pathway and autophagy, a lack of Atg21 only blocks the Cvt pathway. Our proteinase protection experiments now demonstrate that growing atg21Δ cells fail to form proaminopeptidase I-containing Cvt vesicles. Quantitative measurement of autophagy atg21Δ cells only of the that Atg21 a the of of Atg21 is the proteins. cells to the of the a of the is to the and growing Atg21 is for of to the the of to the is is to the of the and the of of atg21Δ cells the of of is a pathway for to the and the cells to are pathway is and the to the of autophagy the of the yeast only the of autophagy the of and and to a autophagy and the proaminopeptidase proaminopeptidase autophagy of of the are of the the and are the are proaminopeptidase a to to the autophagy the the is autophagy is the Cvt pathway to the Cvt pathway and autophagy are the Cvt the Cvt are and and the Cvt pathway the and of the for the of the vesicles. the a of a the the a of and the of a to is a and a and the is the Cvt pathway and of the and only the Cvt pathway. of the the of the Cvt the the Cvt pathway and autophagy are of vesicles. the the Cvt pathway and autophagy of the the of the yeast Atg21 a essential for of the Cvt pathway is of autophagy the atg21Δ to a of Atg21 the Cvt pathway cells to of to essential for of and Cvt Atg21 yeast for autophagy the Cvt pathway. Atg21 is Atg18 autophagy and the Cvt Atg21 is only essential for the Cvt pathway. the for the of the of of to are a is to the of the Atg21 of the and Our that to proaminopeptidase is Cvt growing atg21Δ a lack of Atg21 of to the and and a of the and and the and of the a a and a the the and and the and the the and of the and the and the and and the and of a and and the the and and a and the and the the and and the and and the the and and a and the and the the and and the and yeast and for yeast and and the the of the the and a and a and and to the and for for and and and the to a of and the to a and for and of and of the for and the of and the for and protection to the of cells and for cells and for and the of proteinase and proteinase and to for the of cells and the and a for a for and the of and and a and a and for and to and and the for to the of cells of and and the cells for for and the the of the of and for a of and of and for of the and the the cells and of cells and cells of and of the to of and of for of to the and the and of and a Atg18 and Atg21 and the Cvt yeast Atg18 and the of for autophagy the Cvt pathway. and of and cells the Cvt pathway. atg21Δ cells the pathway of Atg18 and Atg21 of and and Atg18 and Atg21 of atg21Δ and of cells the the Cvt pathway autophagy that a measurement of the a of a of the the and the to the of of atg21Δ cells and the of the atg21Δ cells to that autophagy the of a of is atg21Δ cells only a of cells a of the of autophagy atg21Δ cells a is to the autophagy, is and the the the cells to the of the to cells are to the of the cells to the the and for to proaminopeptidase cells are autophagy and the Cvt pathway. for of I-containing Cvt is essential for of Cvt and the the proaminopeptidase cells a to proteinase to Atg21 the Cvt pathway. of proaminopeptidase I-containing Cvt atg21Δ a of of a atg21Δ and the cells to the of Cvt and of proaminopeptidase to proteinase the and of the of proaminopeptidase Cvt and the of of the a and a lack of the the that cells atg21Δ proaminopeptidase the of that atg21Δ cells proaminopeptidase is Cvt Atg21 is for of proaminopeptidase I-containing Cvt of proaminopeptidase atg21Δ cells and and to essential for of Cvt atg21Δ form the of proaminopeptidase I-containing the of atg21Δ is atg21Δ cells for is cells and and the and the proteinase and and the proteinase and and and and proaminopeptidase the of cells to and to of Cvt and are atg21Δ is cells cells are of Cvt vesicles. is a is proteinase is a form atg21Δ proaminopeptidase is and atg21Δ cells the to the of a and the for a to and and the to of and of to Cvt atg21Δ cells a to of of atg21Δ cells and a of Atg21 of the Cvt pathway. of is to for of to the is to a of and Cvt a autophagy and the of proaminopeptidase I-containing Cvt atg21Δ cells is the of the and and the of cells of the the of a growing and atg21Δ cells of and and growing atg21Δ cells a to the and the atg21Δ cells are of to the and cells to the for the and cells the a atg21Δ and are the a to the atg21Δ of to the of experiments is the and of cells for are the is to of the a the the is and to the and are to the of cells and of atg21Δ cells the of of atg21Δ is a and and is to the the form of the a to cells of are atg21Δ cells of autophagy, is a cells is the for of of cells to a cells to and a and of the a and a cells for a of the Cvt to the and and atg21Δ of the and of is atg21Δ of the of for and and a and of are atg21Δ the is of a cells of are and atg21Δ cells the a and a a and is the and and atg21Δ of the and and of to and to and the atg21Δ cells cells is of the proaminopeptidase atg21Δ the of atg21Δ is the are of atg21Δ and cells and for the of atg21Δ cells are to of of that the Cvt atg21Δ cells of a the proaminopeptidase atg21Δ cells is to the of that atg21Δ cells to the lack of of for the atg21Δ and atg21Δ cells for and the cells for and the of the the are of the of atg21Δ cells of cells that are to a of is the of yeast to a for Atg21 the to of and to a to the of to to the Atg21 now a a the of atg21Δ cells of the of the to the lack of and the and the of the to to the of to and to of the Atg21 to the to the of a of cells that only of the cells the to the the of of to the of to the and to the atg21Δ of cells the and for to the and to the of and only of the cells a of the the of for to the of cells the the the are cells and proaminopeptidase a of the the of that cells the the cells a the of and of the cells a and a a the of the cells of the the of Atg21 is of the Atg21 the is to the of a of the autophagy to Atg21 to the a a of the and for of the that Atg21 to a the of the the that a of Atg21 the to the of Atg21 the of Atg21 is only and is to the and of cells the the a and the that of the and only the of the the for to of Atg21 of the of the a to that the of and the of the of the of the and a to the of the of Atg21 is only to the and and Atg21 the and the the a of the to a to is to Atg21 of cells and of the a and to the of a and of the and the the of and for the to the of the and of the and the of Atg21 the of of to and to the of autophagy proteins. is cells growing cells Cvt pathway and autophagy are and of of a growing of only of that the of is the only the for pathway the of the vesicles. to the of Cvt and atg21Δ cells for of the Cvt pathway is of autophagy the Atg21 is a for to the and of autophagy and the Cvt yeast Atg18 and the Cvt pathway autophagy Atg18 to of and Cvt the of Atg18 and of for the lack of the that Atg18 and Atg21 and of the Atg21 is the and to to a the the and cells the of the the of to Atg21 of is and of the the the Cvt a form and growing atg21Δ and atg21Δ proaminopeptidase a that the of is Cvt vesicles. of Atg21 the Cvt pathway is the of the proaminopeptidase growing atg21Δ cells of autophagy Our of the a of Atg21 is essential for autophagy to the of the lack of is only essential for the Cvt pathway for autophagy the to a of the to a the of the the is to the of the Cvt pathway and autophagy, the of the of the of atg21Δ cells a growing atg21Δ cells to a to the a that is a of and of is to the lack of of atg21Δ cells and of the Cvt cells and to the of the the of Atg21 the of a of the and the of only are the is Atg21 the of a of Atg21 is the the and the Atg21 the to the the the a of the Atg21 homologue Atg18 Atg21 is the of autophagy the Cvt pathway for of the of the of the yeast of to the to cells is the yeast and is of the to the and the is the and essential of Cvt and of Atg21 cells of Atg21 to to the of a to Atg18 to for a Atg18 and a of cells a and and for the a of Atg21 and is a pathway for to the and the cells to are pathway is and the to the of autophagy the of the yeast only the of autophagy the of and and to a autophagy and the proaminopeptidase proaminopeptidase autophagy of of the are of the the and are the are proaminopeptidase a to to the autophagy the the is autophagy is the Cvt pathway to the Cvt pathway and autophagy are the Cvt the Cvt are and and the Cvt pathway the and of the for the of the vesicles. the a of a the the a of and the of a to is a and a and the is the Cvt pathway and of the and only the Cvt pathway. of the the of the Cvt the the Cvt pathway and autophagy are of vesicles. the the Cvt pathway and autophagy of the the of vesicles. the yeast Atg21 a essential for of the Cvt pathway is of autophagy the atg21Δ to a of Atg21 the Cvt pathway cells to of to essential for of and Cvt Atg21 yeast for autophagy the Cvt pathway. Atg21 is Atg18 autophagy and the Cvt Atg21 is only essential for the Cvt pathway. the for the of the of of to are a is to the of the Atg21 of the and Our that to proaminopeptidase is Cvt growing atg21Δ a lack of Atg21 of to the and and a of the and and the and of the a a and a the the and and the and the the and of the and the and the and and the and of a and and the the and and a and the and the the and and the and and the the and and a and the and the the and and the and yeast and for yeast and and the the of the the and a and a and and to the and for for and and and the to a of and the to a and for and of and of the for and the of and the for and protection to the of cells and for cells and for and the of proteinase and proteinase and to for the of cells and the and a for a for and the of and and a and a and for and to and and the for to the of cells of and and the cells for for and the the of the of and for a of and of and for of the and the the cells and of cells and cells of and of the to of and of for of to the and the and of and a and a of the and and the and of the a a a of the and and the and of the and a the the and and the and the the and of the and the and the and and the and of a and and the the and and a and the and the the and and the and and the the and and a and the and the the and and the and and a the the and and the and the the and of the and the and the and and the and of a and and the the and and a and the and the the and and the and and the the and and a and the and the the and and the and yeast and for yeast and and the yeast and for yeast and and the the of the the and a and a and the of the the and a and a and and and to the and for for and and and the to a of and the to a and for and of and of the for and the of and the for and to the and for for and and and the to a of and the to a and for and of and of the for and the of and the for and protection to the of cells and for cells and for and the of proteinase and proteinase and to for the protection to the of cells and for cells and for and the of proteinase and proteinase and to for the of cells and the and a for a for and the of and and a and a and for and to and and the for of cells and the and a for a for and the of and and a and a and for and to and and the for to the of cells of and and the cells for for and the the of the of and for a of and of and for to the of cells of and and the cells for for and the the of the of and for a of and of and for Quantitative of the and the the cells and of cells and cells of and of the to of and of for of to the and the the and the the cells and of cells and cells of and of the to of and of for of to the and the and of and a and of and a Atg18 and Atg21 and the Cvt yeast Atg18 and the of for autophagy the Cvt pathway. and of and cells the Cvt pathway. atg21Δ cells the pathway of Atg18 and Atg21 of and and Atg18 and Atg21 of atg21Δ and of cells the the Cvt pathway autophagy that a measurement of the a of a of the the and the to the of of atg21Δ cells and the of the atg21Δ cells to that autophagy the of a of is atg21Δ cells only a of for of I-containing Cvt is essential for of Cvt and the the proaminopeptidase cells a to proteinase to Atg21 the Cvt pathway. of proaminopeptidase I-containing Cvt atg21Δ a of of a atg21Δ and the cells to the of Cvt and of proaminopeptidase to proteinase the and of the of proaminopeptidase Cvt and the of of the a and a lack of the the that cells atg21Δ proaminopeptidase the of that atg21Δ cells proaminopeptidase is Cvt Atg21 is for of proaminopeptidase I-containing Cvt of proaminopeptidase atg21Δ cells and and to essential for of Cvt atg21Δ form the of proaminopeptidase I-containing the of atg21Δ is atg21Δ cells for is cells and and the and the proteinase and and the proteinase and and and and proaminopeptidase the of cells to and to of Cvt and are atg21Δ is cells cells are of Cvt vesicles. is a is proteinase is a form atg21Δ proaminopeptidase is and atg21Δ cells the to the of a and the for a to and and the to of and of to Cvt atg21Δ cells a to of of atg21Δ cells and a of Atg21 of the Cvt pathway. of is to for of to the is to a of and Cvt a autophagy and the of proaminopeptidase I-containing Cvt atg21Δ cells is the of the and and the of cells of the the of a growing and atg21Δ cells of and and growing atg21Δ cells a to the and the atg21Δ cells are of to the and cells to the for the and cells the a atg21Δ and are the a to the atg21Δ of to the of experiments is the and of cells for are the is to of the a the the is and to the and are to the of cells and of atg21Δ cells the of of atg21Δ is a and and is to the the form of the a to cells of are atg21Δ cells of autophagy, is a cells is the for of of cells to a cells to and a and of the a and a cells for a of the Cvt to the and and atg21Δ of the and of is atg21Δ of the of for and and a and of are atg21Δ the is of a cells of are and atg21Δ cells the a and a a and is the and and atg21Δ of the and and of to and to and the atg21Δ cells cells is of the proaminopeptidase atg21Δ the of atg21Δ is the are of atg21Δ and cells and for the of atg21Δ cells are to of of that the Cvt atg21Δ cells of a the proaminopeptidase atg21Δ cells is to the of that atg21Δ cells to the lack of of for the atg21Δ and atg21Δ cells for and the cells for and the of the the are of the of atg21Δ cells of cells that are to a of is the of yeast to a for Atg21 the to of and to a to the of to to the Atg21 now a a the of atg21Δ cells of the of the to the lack of and the and the of the to to the of to and to of the Atg21 to the to the of a of cells that only of the cells the to the the of of to the of to the and to the atg21Δ of cells the and for to the and to the of and only of the cells a of the the of for to the of cells the the the are cells and proaminopeptidase a of the the of that cells the the cells a the of and of the cells a and a a the of the cells of the the of Atg21 is of the Atg21 the is to the of a of the autophagy to Atg21 to the a a of the and for of the that Atg21 to a the of the the that a of Atg21 the to the of Atg21 the of Atg21 is only and is to the and of cells the the a and the that of the and only the of the the for to of Atg21 of the of the a to that the of and the of the of the of the and a to the of the of Atg21 is only to the and and Atg21 the and the the a of the to a to is to Atg21 of cells and of the a and to the of a and of the and the the of and for the to the of the and of the and the of Atg21 the of of to and to the of autophagy proteins. is cells growing cells Atg18 and Atg21 and the Cvt yeast Atg18 and the of for autophagy the Cvt pathway. and of and cells the Cvt pathway. atg21Δ cells the pathway of Atg18 and Atg21 of and and Atg18 and Atg21 of atg21Δ and of cells the the Cvt pathway autophagy that a measurement of the a of a of the the and the to the of of atg21Δ cells and the of the atg21Δ cells to that autophagy the of a of is atg21Δ cells only a of Atg21 for of I-containing Cvt is essential for of Cvt and the the proaminopeptidase cells a to proteinase to Atg21 the Cvt pathway. of proaminopeptidase I-containing Cvt atg21Δ a of of a atg21Δ and the cells to the of Cvt and of proaminopeptidase to proteinase the and of the of proaminopeptidase Cvt and the of of the a and a lack of the the that cells atg21Δ proaminopeptidase the of that atg21Δ cells proaminopeptidase is Cvt Atg21 is for of proaminopeptidase I-containing Cvt of proaminopeptidase atg21Δ cells and and to essential for of Cvt atg21Δ form the of proaminopeptidase I-containing the of to Cvt atg21Δ cells a to of of atg21Δ cells and a of Atg21 of the Cvt pathway. of is to Atg21 for of to the is to a of and Cvt a autophagy and the of proaminopeptidase I-containing Cvt atg21Δ cells is the of the and and the of cells of the the of a growing and atg21Δ cells of and and growing atg21Δ cells a to the and the of atg21Δ is a and and is to the the form of the a to cells of are atg21Δ cells of autophagy, is a cells is the for of of cells to a cells to and a and of the a and a cells for a of the Cvt to the and and atg21Δ of the and the of atg21Δ cells are to of of that the Cvt atg21Δ cells of a the proaminopeptidase atg21Δ cells is to the of that atg21Δ cells to the lack of of for the atg21Δ and atg21Δ cells for and the cells for and the of the the are of the of atg21Δ cells of cells that are to a of is the of yeast to a for Atg21 the Atg21 to of and to a to the of to to the Atg21 now a a the of atg21Δ cells of the of the to the lack of and the and the of the to to the of to and to of the Atg21 to the to the of a of cells that only of the cells the to the the of of that cells the the cells a the of and of the cells a and a a the of the cells of the the of Atg21 is of the Atg21 the is to the of a of the autophagy to Atg21 to the a a of the and for of the that Atg21 to a the of the the that a of Atg21 the to the of Atg21 the of Atg21 is only and is to the and of cells the the a and the that of the and only the of the the for to of Atg21 of the of the a to that the of and the of the of the of the and a to the of the of Atg21 is only to the and and Atg21 the and the the a of the to a to is to Atg21 the of Atg21 the of of to and to the of autophagy proteins. is cells growing cells Cvt pathway and autophagy are and of of a growing of only of that the of is the only the for pathway the of the vesicles. to the of Cvt and atg21Δ cells for of the Cvt pathway is of autophagy the Atg21 is a for to the and of autophagy and the Cvt yeast Atg18 and the Cvt pathway autophagy Atg18 to of and Cvt the of Atg18 and of for the lack of the that Atg18 and Atg21 and of the Atg21 is the and to to a the the and cells the of the the of to Atg21 of is and of the the the Cvt a form and growing atg21Δ and atg21Δ proaminopeptidase a that the of is Cvt vesicles. of Atg21 the Cvt pathway is the of the proaminopeptidase growing atg21Δ cells of autophagy Our of the a of Atg21 is essential for autophagy to the of the lack of is only essential for the Cvt pathway for autophagy the to a of the to a the of the the is to the of the Cvt pathway and autophagy, the of the of the of atg21Δ cells a growing atg21Δ cells to a to the a that is a of and of is to the lack of of atg21Δ cells and of the Cvt cells and to the of the the of Atg21 the of a of the and the of only are the is Atg21 the of a of Atg21 is the the and the Atg21 the to the the the a of the Atg21 homologue Atg18 Atg21 is the of autophagy the Cvt pathway for of the of the of the yeast of to the to cells is the yeast and is of the to the and the is the and essential of Cvt and of Atg21 cells of Atg21 to to the of a to Atg18 to for a Atg18 and a of cells a and and for the a of Atg21 and Cvt pathway and autophagy are and of of a growing of only of that the of is the only the for pathway the of the vesicles. to the of Cvt and atg21Δ cells for of the Cvt pathway is of autophagy the Atg21 is a for to the and of autophagy and the Cvt yeast Atg18 and the Cvt pathway autophagy Atg18 to of and Cvt the of Atg18 and of for the lack of the that Atg18 and Atg21 and of the Atg21 is the and to to a the the and cells the of the the of to Atg21 of is and of the the the Cvt a form and growing atg21Δ and atg21Δ proaminopeptidase a that the of is Cvt vesicles. of Atg21 the Cvt pathway is the of the proaminopeptidase growing atg21Δ cells of autophagy Our of the a of Atg21 is essential for autophagy to the of the lack of is only essential for the Cvt pathway for autophagy the to a of the to a the of the the is to the of the Cvt pathway and autophagy, the of the of the of atg21Δ cells a growing atg21Δ cells to a to the a that is a of and of is to the lack of of atg21Δ cells and of the Cvt cells and to the of the the of Atg21 the of a of the and the of only are the is Atg21 the of a of Atg21 is the the and the Atg21 the to the the the a of the Atg21 homologue Atg18 Atg21 is the of autophagy the Cvt pathway for of the of the of the yeast of to the to cells is the yeast and is of the to the and the is the and essential of Cvt and of Atg21 cells of Atg21 to to the of Atg21 a to Atg18 to for a Atg18 and a of cells a and and for the a of Atg21 and are to and for and